NleC, a type III secretion protease, compromises NF-κB activation by targeting p65/RelA

Area of Science:

• Microbiology
• Immunology
• Molecular Biology

Background:

• The NF-κB signaling pathway is crucial for immune responses, orchestrating the production of inflammatory cytokines.
• Pathogens like enteropathogenic and enterohemorrhagic Escherichia coli (EPEC/EHEC) have evolved mechanisms to evade host immunity by subverting the NF-κB pathway.
• EPEC/EHEC utilize the type three secretion system (T3SS) to inject bacterial effectors that manipulate host cell functions, including immune suppression.

Purpose of the Study:

• To identify bacterial factors employed by EPEC/EHEC to suppress the NF-κB signaling pathway and inhibit inflammatory responses.
• To elucidate the molecular mechanisms by which these effectors interfere with NF-κB activation.
• To assess the contribution of individual and combined effectors to the overall immune suppressive capacity of EPEC/EHEC.

Main Methods:

• Utilized gain- and loss-of-function screenings to identify bacterial effectors involved in NF-κB regulation.
• Investigated the enzymatic activity of identified effectors, specifically characterizing NleC as a protease.
• Analyzed the impact of single and double effector knockouts (ΔnleC, ΔnleE, ΔnleC ΔnleE) on NF-κB activation and cytokine production in host cells.

Main Results:

• Identified NleC and NleE as key bacterial effectors that down-regulate NF-κB signaling upon T3SS-mediated injection.
• Demonstrated that NleC is a zinc protease that directly cleaves the NF-κB p65 subunit, reducing its nuclear translocation and activity.
• Showed that the combined absence of NleC and NleE (ΔnleC ΔnleE) resulted in a significantly enhanced inflammatory response, comparable to a T3SS-defective mutant, indicating their cooperative role in immune suppression.

Conclusions:

• NleC functions as an anti-inflammatory bacterial zinc protease that disrupts NF-κB activation through p65 cleavage.
• The cooperative action of NleC and NleE is essential for the efficient suppression of the NF-κB pathway by EPEC/EHEC.
• These effectors collectively account for the majority of immune suppression mediated by EHEC/EPEC, highlighting a sophisticated bacterial strategy to evade host defenses.